Alpha-fetoprotein (AFP) lectin-binding molecular variants synthesized by a series of transplantable rat hepatomas, each hepatoma line secreting a characteristic "mutant" pattern of variants, will serve as a model for glycosylation alterations associated with malignancy. Initial assessment of the degree and type of glycan heterogeneity in tumor-produced AFP will be made by lectin crossed-immunoaffinoelectrophoresis. To identify specific changes or classes of changes in glycosylation, we will determine the primary structure of the AFP secreted by normal hepatocytes (newborn liver) and by malignant hepatocytes (transplantable hepatomas). This will show whether or not the altered patterns of lectin-binding molecular variants secreted by malignant hepatocytes reflect abnormal proportions of AFP molecules with a normal oligosaccharide composition or characteristic proportions of AFP molecules with altered oligosaccharide composition. Molar ratios of monosaccharides will be estimated by gas-liquid chromatography; glycan structure by thin-layer chromatography and paper electrophoresis; linkages between monosaccharides by methylation analysis and mass spectrometry and nuclear magnetic resonance spectroscopy. Elucidation of the carbohydrate structures of the AFP lectin-binding variants also will provide information on the homogeneity or heterogeneity of glycosylation systems, which is relevant to the synthesis of carbohydrate units in normal cells. Similar studies are being carried out on glycosylation of human AFP during newborn growth, nonmalignant liver disease and by primary hepatocellular carcinoma. Patterns of AFP glycosylation abnormalities parallel those demonstrated in rats. Analysis of AFP lectinaffinity patterns may improve discrimination between malignant and nonmalignant liver disease when serum AFP concentrations are not diagnostic of hepatoma. AFP lectin-binding properties may provide a marker for the transition from premalignant liver disease to malignancy and a means to detect small resectable tumors earlier.